ABSTRACT
The commitment of stem cells to differentiate into osteoblasts is a highly regulated and complex process that involves the coordination of extrinsic signals and intrinsic transcriptional machinery. While rodent osteoblastic differentiation has been extensively studied, research on human osteogenesis has been limited by cell sources and existing models. Here, we systematically dissect human pluripotent stem cell-derived osteoblasts to identify functional membrane proteins and their downstream transcriptional networks involved in human osteogenesis. Our results reveal an enrichment of type II transmembrane serine protease CORIN in humans but not rodent osteoblasts. Functional analyses demonstrated that CORIN depletion significantly impairs osteogenesis. Genome-wide chromatin immunoprecipitation enrichment and mechanistic studies show that p38 MAPK-mediated CCAAT enhancer binding protein delta (CEBPD) upregulation is required for CORIN-modulated osteogenesis. Contrastingly, the type I transmembrane heparan sulfate proteoglycan SDC1 enriched in mesenchymal stem cells exerts a negative regulatory effect on osteogenesis through a similar mechanism. Chromatin immunoprecipitation-seq, bulk and single-cell transcriptomes, and functional validations indicated that CEBPD plays a critical role in controlling osteogenesis. In summary, our findings uncover previously unrecognized CORIN-mediated CEBPD transcriptomic networks in driving human osteoblast lineage commitment.
ABSTRACT
BACKGROUND: Cell-free DNA (cfDNA) is being explored as biomarker for non-invasive diagnosis of cancer. We aimed to establish a cfDNA-based DNA methylation marker panel to differentially diagnose papillary thyroid carcinoma (PTC) from benign thyroid nodule (BTN). METHODS: 220 PTC- and 188 BTN patients were enrolled. Methylation markers of PTC were identified from patients' tissue and plasma by reduced representation bisulfite sequencing and methylation haplotype analyses. They were combined with PTC markers from literatures and were tested on additional PTC and BTN samples to verify PTC-detecting ability using targeted methylation sequencing. Top markers were developed into ThyMet and were tested in 113 PTC and 88 BTN cases to train and validate a PTC-plasma classifier. Integration of ThyMet and thyroid ultrasonography was explored to improve accuracy. FINDINGS: From 859 potential PTC plasma-discriminating markers that include 81 markers identified by us, the top 98 most PTC plasma-discriminating markers were selected for ThyMet. A 6-marker ThyMet classifier for PTC plasma was trained. In validation it achieved an Area Under the Curve (AUC) of 0.828, similar to thyroid ultrasonography (0.833) but at higher specificity (0.722 and 0.625 for ThyMet and ultrasonography, respectively). A combinatorial classifier by them, ThyMet-US, improved AUC to 0.923 (sensitivity = 0.957, specificity = 0.708). INTERPRETATION: The ThyMet classifier improved the specificity of differentiating PTC from BTN over ultrasonography. The combinatorial ThyMet-US classifier may be effective in preoperative diagnosis of PTC. FUNDING: This work was supported by the grants from National Natural Science Foundation of China (82072956 and 81772850).
Subject(s)
Thyroid Neoplasms , Thyroid Nodule , Humans , Thyroid Cancer, Papillary/diagnosis , Thyroid Cancer, Papillary/genetics , Thyroid Nodule/diagnosis , Thyroid Nodule/genetics , Thyroid Nodule/pathology , DNA Methylation , Thyroid Neoplasms/diagnosis , Thyroid Neoplasms/genetics , Thyroid Neoplasms/pathology , Biomarkers , Biomarkers, Tumor/metabolismABSTRACT
Amyotrophic lateral sclerosis (ALS) is a progressive neurodegenerative disease that results in the loss of motor function in the central nervous system (CNS) and ultimately death. The mechanisms underlying ALS pathogenesis have not yet been fully elucidated, and ALS cannot be treated effectively. Most studies have applied animal or single-gene intervention cell lines as ALS disease models, but they cannot accurately reflect the pathological characteristics of ALS. Induced pluripotent stem cells (iPSCs) can be reprogrammed from somatic cells, possessing the ability to self-renew and differentiate into a variety of cells. iPSCs can be obtained from ALS patients with different genotypes and phenotypes, and the genetic background of the donor cells remains unchanged during reprogramming. iPSCs can differentiate into neurons and glial cells related to ALS. Therefore, iPSCs provide an excellent method to evaluate the impact of diseases on ALS patients. Moreover, patient-derived iPSCs are obtained from their own somatic cells, avoiding ethical concerns and posing only a low risk of immune rejection. The iPSC technology creates new hope for ALS treatment. Here, we review recent studies on iPSCs and their applications in disease modeling, drug screening and cell therapy in ALS, with a particular focus on the potential for ALS treatment.
Subject(s)
Amyotrophic Lateral Sclerosis , Induced Pluripotent Stem Cells , Neurodegenerative Diseases , Animals , Amyotrophic Lateral Sclerosis/genetics , Amyotrophic Lateral Sclerosis/therapy , Amyotrophic Lateral Sclerosis/metabolism , Induced Pluripotent Stem Cells/metabolism , Neurodegenerative Diseases/metabolism , Neurons/metabolism , Cell- and Tissue-Based TherapyABSTRACT
BACKGROUND: Metastasis has emerged to be an important cause for poor prognosis of thyroid carcinoma (TC) and its molecular mechanisms are not fully understood. STRA6 is a multifunctional membrane protein widely expressed in embryonic and adult tissues. The function and mechanism of STRA6 in TC remain elusive. OBJECTIVE: We aimed to explore the role of STRA6 in TC progression and provide a therapeutic target for TC. METHODS: The expression and clinicopathological relevance of STRA6 were explored in TC. Stable STRA6-knockdown TC cells were established and used to determine the biological function of STRA6 in vitro and in vivo. RNA sequencing and co-immunoprecipitation were performed to unveil the molecular mechanism of STRA6 in TC progression. The potential of STRA6 as a therapeutic target was evaluated by lipid nanoparticles (LNPs) containing siRNA. RESULTS: STRA6 was upregulated in TC and correlated with aggressive clinicopathological features, including extrathyroidal extension and lymph node metastasis, which contributed to the poor prognosis of TC. STRA6 facilitated TC progression by enhancing proliferation and metastasis in vitro and in vivo. Mechanistically, STRA6 could interact with integrin-linked kinase (ILK) and subsequently activate the protein kinase B/mechanistic target of rapamycin (AKT/mTOR) signaling pathway. We further unveiled that STRA6 reprogrammed lipid metabolism through SREBP1, which was crucial for the metastasis of TC. Moreover, STRA6 siRNA delivered by LNPs significantly inhibited cell growth in xenograft tumor models. CONCLUSIONS: Our study demonstrates the critical roles of STRA6 contributing to TC progression via the ILK/AKT/mTOR axis, which may provide a novel prognostic marker as well as a promising therapeutic target for aggressive TC.
Subject(s)
Proto-Oncogene Proteins c-akt , Thyroid Neoplasms , Animals , Mice , Humans , Female , Proto-Oncogene Proteins c-akt/metabolism , Mice, Nude , TOR Serine-Threonine Kinases/metabolism , Thyroid Neoplasms/genetics , RNA, Small Interfering , Cell Line, Tumor , Cell Proliferation/genetics , Disease Models, Animal , Membrane Proteins/geneticsABSTRACT
CONTEXT: Both Graves disease (GD) and inflammatory bowel disease (IBD) are common autoimmune diseases that severely damage a patient's quality of life. Previous epidemiological studies have suggested associations between GD and IBD. However, whether a causal relationship exists between these 2 diseases remains unknown. OBJECTIVE: To infer a causal relationship between GD and IBD using bidirectional 2-sample Mendelian randomization (MR). METHODS: We performed bidirectional 2-sample MR to infer a causal relationship between GD and IBD using genome-wide association study summary data obtained from Biobank Japan and the International Inflammatory Bowel Disease Genetic Consortium. Several methods (random-effect inverse variance weighted, weighted median, MR-Egger regression, and MR-PRESSO) were used to ensure the robustness of the causal effect. Heterogeneity was measured based on Cochran's Q value. Horizontal pleiotropy was evaluated by MR-Egger regression and leave-one-out analysis. RESULTS: Genetically predicted IBD may increase the risk of GD by 24% (odds ratio [OR] 1.24, 95% CI 1.01-1.52, P = .041). Crohn disease (CD) may increase the risk of GD, whereas ulcerative colitis (UC) may prevent patients from developing GD. Conversely, genetically predicted GD may slightly increase the risk of CD, although evidence indicating that the presence of GD increased the risk of UC or IBD was lacking. Outlier-corrected results were consistent with raw causal estimates. CONCLUSION: Our study revealed a potentially higher comorbidity rate for GD and CD. However, UC might represent a protective factor for GD. The underlying mechanism and potential common pathways await discovery.
Subject(s)
Colitis, Ulcerative , Crohn Disease , Graves Disease , Inflammatory Bowel Diseases , Humans , Genome-Wide Association Study , Mendelian Randomization Analysis , Quality of Life , Inflammatory Bowel Diseases/epidemiology , Inflammatory Bowel Diseases/genetics , Colitis, Ulcerative/epidemiology , Colitis, Ulcerative/genetics , Crohn Disease/epidemiology , Crohn Disease/genetics , Graves Disease/epidemiology , Graves Disease/geneticsABSTRACT
Osteosarcoma (OS) is a highly malignant tumor occurring in bone tissue with a high propensity to metastasize, and its underlying mechanisms remain largely elusive. The OS prognosis is poor, and improving the survival of OS patients remains a challenge. Current treatment methods such as surgical approaches, chemotherapeutic drugs, and immunotherapeutic drugs remain ineffective. As research progresses, targeted therapy is gradually becoming irreplaceable. In this review, several treatment modalities for osteosarcoma, such as surgery, chemotherapy, and immunotherapy, are briefly described, followed by a discussion of targeted therapy, the important targets, and new technologies for osteosarcoma treatment.
Subject(s)
Bone Neoplasms , Osteosarcoma , Humans , Bone Neoplasms/pathology , Osteosarcoma/pathology , Molecular Targeted Therapy , Bone and Bones/pathology , Immunologic Factors/therapeutic useABSTRACT
Background: Several epidemiological studies have reported a possible correlation between risk of gout and metabolic disorders including type 2 diabetes, insulin resistance, obesity, dyslipidemia, and hypertension. However, it is unclear if this association is causal. Methods: We used Mendelian randomization (MR) to evaluate the causal relation between metabolic conditions and gout or serum urate concentration by inverse-variance-weighted (conventional) and weighted median methods. Furthermore, MR-Egger regression and MR-pleiotropy residual sum and outlier (PRESSO) method were used to explore pleiotropy. Genetic instruments for metabolic disorders and outcome (gout and serum urate) were obtained from several genome-wide association studies on individuals of mainly European ancestry. Results: Conventional MR analysis showed a robust causal association of increasing obesity measured by body mass index (BMI), high-density lipoprotein cholesterol (HDL), and systolic blood pressure (SBP) with risk of gout. A causal relationship between fasting insulin, BMI, HDL, triglycerides (TG), SBP, alanine aminotransferase (ALT), and serum urate was also observed. These results were consistent in weighted median method and MR-PRESSO after removing outliers identified. Our analysis also indicated that HDL and serum urate as well as gout have a bidirectional causal effect on each other. Conclusions: Our study suggested causal effects between glycemic traits, obesity, dyslipidemia, blood pressure, liver function, and serum urate as well as gout, which implies that metabolic factors contribute to the development of gout via serum urate, as well as potential benefit of sound management of increased serum urate in patients with obesity, dyslipidemia, hypertension, and liver dysfunction.
Subject(s)
Diabetes Mellitus, Type 2 , Gout , Hypertension , Diabetes Mellitus, Type 2/epidemiology , Diabetes Mellitus, Type 2/genetics , Genome-Wide Association Study , Gout/epidemiology , Gout/genetics , Humans , Mendelian Randomization Analysis/methods , Obesity/complications , Obesity/epidemiology , Obesity/genetics , Uric AcidABSTRACT
Amyotrophic lateral sclerosis (ALS) is a rapidly progressive and highly fatal neurodegenerative disease. Although the pathogenesis of ALS remains unclear, increasing evidence suggests that a key contributing factor is mitochondrial dysfunction. Mitochondria are organelles in eukaryotic cells responsible for bioenergy production, cellular metabolism, signal transduction, calcium homeostasis, and immune responses and the stability of their function plays a crucial role in neurons. A single disorder or defect in mitochondrial function can lead to pathological changes in cells, such as an impaired calcium buffer period, excessive generation of free radicals, increased mitochondrial membrane permeability, and oxidative stress (OS). Recent research has also shown that these mitochondrial dysfunctions are also associated with pathological changes in ALS and are believed to be commonly involved in the pathogenesis of the disease. This article reviews the latest research on mitochondrial dysfunction and its impact on the progression of ALS, with specific attention to the potential of novel therapeutic strategies targeting mitochondrial dysfunction.
Subject(s)
Amyotrophic Lateral Sclerosis , Neurodegenerative Diseases , Amyotrophic Lateral Sclerosis/metabolism , Calcium/metabolism , Humans , Mitochondria/metabolism , Neurodegenerative Diseases/metabolism , Oxidative Stress/physiologyABSTRACT
The RB1 gene is frequently mutated in human cancers but its role in tumorigenesis remains incompletely defined. Using an induced pluripotent stem cell (iPSC) model of hereditary retinoblastoma (RB), we report that the spliceosome is an up-regulated target responding to oncogenic stress in RB1-mutant cells. By investigating transcriptomes and genome occupancies in RB iPSCderived osteoblasts (OBs), we discover that both E2F3a, which mediates spliceosomal gene expression, and pRB, which antagonizes E2F3a, coregulate more than one-third of spliceosomal genes by cobinding to their promoters or enhancers. Pharmacological inhibition of the spliceosome in RB1-mutant cells leads to global intron retention, decreased cell proliferation, and impaired tumorigenesis. Tumor specimen studies and genome-wide TCGA (The Cancer Genome Atlas) expression profile analyses support the clinical relevance of pRB and E2F3a in modulating spliceosomal gene expression in multiple cancer types including osteosarcoma (OS). High levels of pRB/E2F3aregulated spliceosomal genes are associated with poor OS patient survival. Collectively, these findings reveal an undiscovered connection between pRB, E2F3a, the spliceosome, and tumorigenesis, pointing to the spliceosomal machinery as a potentially widespread therapeutic vulnerability of pRB-deficient cancers.
Subject(s)
Bone Neoplasms , Carcinogenesis , E2F3 Transcription Factor , Gene Expression Regulation, Neoplastic , Induced Pluripotent Stem Cells , Osteosarcoma , Retinoblastoma Binding Proteins , Spliceosomes , Ubiquitin-Protein Ligases , Bone Neoplasms/genetics , Bone Neoplasms/pathology , Carcinogenesis/genetics , E2F3 Transcription Factor/genetics , E2F3 Transcription Factor/metabolism , Genes, Retinoblastoma , Humans , Induced Pluripotent Stem Cells/metabolism , Mutation , Osteosarcoma/genetics , Osteosarcoma/pathology , Retinal Neoplasms/genetics , Retinoblastoma/genetics , Retinoblastoma Binding Proteins/genetics , Retinoblastoma Binding Proteins/metabolism , Spliceosomes/genetics , Spliceosomes/metabolism , Ubiquitin-Protein Ligases/genetics , Ubiquitin-Protein Ligases/metabolismABSTRACT
Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease that affects upper and lower motor neurons. As there is no effective treatment for ALS, it is particularly important to screen key gene therapy targets. The identifications of microRNAs (miRNAs) have completely changed the traditional view of gene regulation. miRNAs are small noncoding single-stranded RNA molecules involved in the regulation of post-transcriptional gene expression. Recent advances also indicate that miRNAs are biomarkers in many diseases, including neurodegenerative diseases. In this review, we summarize recent advances regarding the mechanisms underlying the role of miRNAs in ALS pathogenesis and its application to gene therapy for ALS. The potential of miRNAs to target diverse pathways opens a new avenue for ALS therapy.
Subject(s)
Amyotrophic Lateral Sclerosis , MicroRNAs , Neurodegenerative Diseases , Amyotrophic Lateral Sclerosis/metabolism , Biomarkers/metabolism , Humans , MicroRNAs/genetics , MicroRNAs/metabolism , Motor Neurons/pathology , Neurodegenerative Diseases/metabolismABSTRACT
Rothmund-Thomson syndrome (RTS) is an autosomal recessive genetic disorder characterized by poikiloderma, small stature, skeletal anomalies, sparse brows/lashes, cataracts, and predisposition to cancer. Type 2 RTS patients with biallelic RECQL4 pathogenic variants have multiple skeletal anomalies and a significantly increased incidence of osteosarcoma. Here, we generated RTS patient-derived induced pluripotent stem cells (iPSCs) to dissect the pathological signaling leading to RTS patient-associated osteosarcoma. RTS iPSC-derived osteoblasts showed defective osteogenic differentiation and gain of in vitro tumorigenic ability. Transcriptome analysis of RTS osteoblasts validated decreased bone morphogenesis while revealing aberrantly upregulated mitochondrial respiratory complex I gene expression. RTS osteoblast metabolic assays demonstrated elevated mitochondrial respiratory complex I function, increased oxidative phosphorylation (OXPHOS), and increased ATP production. Inhibition of mitochondrial respiratory complex I activity by IACS-010759 selectively suppressed cellular respiration and cell proliferation of RTS osteoblasts. Furthermore, systems analysis of IACS-010759-induced changes in RTS osteoblasts revealed that chemical inhibition of mitochondrial respiratory complex I impaired cell proliferation, induced senescence, and decreased MAPK signaling and cell cycle associated genes, but increased H19 and ribosomal protein genes. In summary, our study suggests that mitochondrial respiratory complex I is a potential therapeutic target for RTS-associated osteosarcoma and provides future insights for clinical treatment strategies.
Subject(s)
Electron Transport Complex I/genetics , Osteosarcoma/genetics , RNA, Long Noncoding/genetics , RecQ Helicases/genetics , Rothmund-Thomson Syndrome/genetics , Adenosine Triphosphate/biosynthesis , Cell Proliferation/drug effects , Cell Respiration/drug effects , Cellular Senescence/genetics , Electron Transport Complex I/antagonists & inhibitors , Gene Expression Regulation, Developmental/genetics , Humans , Induced Pluripotent Stem Cells/drug effects , Induced Pluripotent Stem Cells/metabolism , Mitogen-Activated Protein Kinase Kinases/genetics , Mutation/genetics , Osteoblasts/drug effects , Osteogenesis/genetics , Osteosarcoma/complications , Osteosarcoma/pathology , Oxadiazoles/pharmacology , Oxidative Phosphorylation/drug effects , Piperidines/pharmacology , Rothmund-Thomson Syndrome/complications , Rothmund-Thomson Syndrome/pathologyABSTRACT
BACKGROUND: Myeloid-derived suppressor cells (MDSCs) have become increasingly recognized as facilitators of tumor development. However, the role of MDSCs in papillary thyroid carcinoma (PTC) progression has not been clearly explored. OBJECTIVE: We aimed to evaluate the levels and function of circulating MDSCs in PTC. METHODS: The proportion of circulating polymorphonuclear (PMN)-MDSCs and mononuclear-MDSCs from patients with PTC or benign thyroid nodules and healthy controls was measured using flow cytometry. For immunosuppressive activity analysis, sorted circulating MDSCs were cocultured with CD3/CD28-costimulated T lymphocytes and the proliferation of T cells was determined. PTC cell lines (TPC-1 and BC-PAP) were cocultured with PMN-MDSCs, and the effects on cell migration, invasion, proliferation, and apoptosis were evaluated. The differential expressed microribonucleic acids (RNAs) and messenger RNAs and their function were also explored in TPC-1 cells cocultured with or without PMN-MDSCs. RESULTS: PMN-MDSCs were increased in peripheral blood mononuclear cells of patients with PTC. Circulating PMN-MDSCs displayed strong T cell suppressive activity. PTC cells demonstrated enhanced invasive capabilities in vitro and in vivo when cocultured with sorted PMN-MDSCs. PMN-MDSCs decreased expression of miR-486-3p and activated nuclear factor kappa B2 (NF-κB2), a direct target of miR-486-3p. Rescue of miR-486-3p diminished the cell migration and invasion induced by PMN-MDSCs. CONCLUSION: Collectively, our work indicates that circulating PMN-MDSCs promote PTC progression. By suppressing miR-486-3p, PMN-MDSCs promote the activity of the NF-κB2 signaling pathway, resulting in accelerated invasion of PTC cells, which may provide new therapeutic strategies for treatment of thyroid cancer.
Subject(s)
Gene Expression Regulation, Neoplastic/genetics , MicroRNAs/metabolism , Myeloid-Derived Suppressor Cells/metabolism , Thyroid Cancer, Papillary/genetics , Thyroid Neoplasms/genetics , Adolescent , Adult , Aged , Apoptosis/drug effects , Apoptosis/genetics , Cell Culture Techniques , Cell Line, Tumor , Cell Movement/drug effects , Cell Movement/genetics , Cell Proliferation/drug effects , Cell Proliferation/genetics , Coculture Techniques , Disease Progression , Female , Flow Cytometry , Gene Expression Regulation, Neoplastic/drug effects , HEK293 Cells , Humans , MicroRNAs/agonists , MicroRNAs/antagonists & inhibitors , Middle Aged , NF-kappa B p52 Subunit/genetics , NF-kappa B p52 Subunit/metabolism , Neoplasm Invasiveness/genetics , Signal Transduction/drug effects , Signal Transduction/genetics , T-Lymphocytes , Thyroid Cancer, Papillary/blood , Thyroid Cancer, Papillary/pathology , Thyroid Cancer, Papillary/surgery , Thyroid Gland/pathology , Thyroid Gland/surgery , Thyroid Neoplasms/blood , Thyroid Neoplasms/pathology , Thyroidectomy , Young AdultABSTRACT
CRISPR, Clustered Regularly Interspaced Short Palindromic Repeat, as a powerful genome engineering system has been widely accepted and employed in gene editing of a vast range of cell types. Comparing to zinc finger nucleases (ZFNs) or transcription-activator-like effector nucleases (TALENs), CRISPR shows less complicated process and higher efficiency. With the development of different CRISPR systems, it can be used not only to knock out a gene, but also to make precise modifications, activate or repress target genes with epigenetic modifications, and even for genome-wide screening. Here we will describe the procedure of generating stable cell lines with a knock-in mutation created by CRISPR. Specifically, this protocol demonstrated how to apply CRISPR to create the point mutation of R249 to S249 on TP53 exon 7 in human embryonic stem cells (hESC) H9 line, which includes three major steps: (1) design CRISPR system targeting TP53 genomic region, (2) deliver the system to H9 hESC and clone selection, and (3) examination and selection of positive clones.
Subject(s)
CRISPR-Cas Systems , Gene Editing , Gene Targeting , Genetic Engineering , Mutation , Animals , Cell Line , Cloning, Molecular , Gene Order , Genetic Vectors , Humans , Plasmids , RNA, Guide, KinetoplastidaABSTRACT
A multitude of signals are coordinated to maintain self-renewal in embryonic stem cells (ESCs). To unravel the essential internal and external signals required for sustaining the ESC state, we expand upon a set of ESC pluripotency-associated phosphoregulators (PRs) identified previously by short hairpin RNA (shRNA) screening. In addition to the previously described Aurka, we identify 4 additional PRs (Bub1b, Chek1, Ppm1g, and Ppp2r1b) whose depletion compromises self-renewal and leads to consequent differentiation. Global gene expression profiling and computational analyses reveal that knockdown of the 5 PRs leads to DNA damage/genome instability, activating p53 and culminating in ESC differentiation. Similarly, depletion of genome integrity-associated genes involved in DNA replication and checkpoint, mRNA processing, and Charcot-Marie-Tooth disease lead to compromise of ESC self-renewal via an increase in p53 activity. Our studies demonstrate an essential link between genomic integrity and developmental cell fate regulation in ESCs.
Subject(s)
Cell Differentiation/genetics , Embryonic Stem Cells/physiology , Genomic Instability , Animals , Cell Cycle Proteins/genetics , Cell Cycle Proteins/physiology , Cell Line , DNA Damage , Gene Expression Profiling , Genetic Complementation Test , Mice , Phosphoproteins/genetics , Phosphoproteins/physiology , RNA, Small Interfering , Signal Transduction , Tumor Suppressor Protein p53/physiologyABSTRACT
Nasopharyngeal carcinoma (NPC), arising from the nasopharynx epithelium, is prevalent among South and East Asia. The radiotherapy is the primary treatment for NPC patients. However, the acquired radioresistance dramatically diminishes the therapeutic effect of radiotherapy. Meanwhile, recurrence and metastasis always occur in line with the radioresistance, but the underlying mechanisms are still unclear. In this study, we established two radioresistant NPC cell lines, CNE1R and SUNE1R, by sequentially irradiated parental CNE1 and SUNE1 cells up to a clinical treatment dose of 72 Gy. A transcriptome profile analysis of CNE1R and CNE1 reveals that activated oncogenic pathways are highly enriched in CNE1R. As the result, CNE1R showed higher proliferation rate but lower apoptosis rate after irradiation, and enhanced metastasis ability in comparison with CNE1. Significantly, a group of metastasis associated genes were increased in CNE1R while the irradiation proceeded, including several matrix metallopeptidase (MMP) members, especially MMP10 and MMP13. With further analysis, we found both MMP10 and MMP13 are highly upregulated in metastatic head and neck cancer specimens compared to non-metastatic ones. More importantly, patients with lower expression of both MMP10 and MMP13 showed a better five-year survival than the double high group. Our findings unveiled the potential mechanisms of radioresistance related metastasis in NPC patients, and the increase of MMP10 and MMP13 may serve as high risk factors for metastasis during radiotherapy.
ABSTRACT
As one of the most essential genome guardians, p53 and its mutants have been suggested associated with many types of cancers. Many p53 mutants function induce unique phenotypes, including carcinogenesis, metastasis, and drug resistance. The p53(R249S) mutation is the most prevalent and specific mutation associated with liver cancer development. Here, we demonstrate the generation of a heterozygous p53(R249S) mutation in the H9 human embryonic stem cell line using TALEN-mediated genome editing. The generated cell line maintains a normal karyotype, a pluripotent state and the in vivo capacity to develop a teratoma containing all three germ layer tissues.
Subject(s)
Cell Culture Techniques/methods , Gene Editing , Human Embryonic Stem Cells/metabolism , Mutation/genetics , Transcription Activator-Like Effector Nucleases/metabolism , Tumor Suppressor Protein p53/genetics , Animals , Base Sequence , Cell Line , Female , Heterozygote , Humans , MiceABSTRACT
The DNA helicase RECQL4 is known for its roles in DNA replication and repair. RECQL4 mutations cause several genetic disorders including Rothmund-Thomson syndrome (RTS), characterized by developmental defects and predisposition to osteosarcoma. Here we reprogrammed fibroblasts with a heterozygous RECQL4 mutation (c.1878â¯+â¯32_1878â¯+â¯55del24) to induced pluripotent stem cells (iPSCs). These iPSCs are pluripotent and are able to be differentiated into all three germ layers, providing a novel tool to further interrogate the role of RECQL4 DNA helicase in vitro.
Subject(s)
Induced Pluripotent Stem Cells/metabolism , RecQ Helicases/genetics , Adult , Female , Heterozygote , Humans , Mutation , Young AdultABSTRACT
Genetic mutations in TP53 contribute to multiple human cancers. Here we report the generation of a H1-p53(R248W/R248W) human embryonic stem cell line harboring a homozygous TP53 R248W mutation created by TALEN-mediated precise gene editing. The H1-p53(R248W/R248W) cell line maintains a normal karyotype, robust pluripotency gene expression, and the potential to differentiate to the three germ layers.
Subject(s)
Gene Editing , Homozygote , Human Embryonic Stem Cells/metabolism , Mutation , Transcription Activator-Like Effector Nucleases , Tumor Suppressor Protein p53/genetics , Cell Line , Gene Expression Regulation , Human Embryonic Stem Cells/cytology , Humans , Male , Tumor Suppressor Protein p53/metabolismABSTRACT
BACKGROUND: We aimed to investigate the roles of hemoglobin (Hb) concentrations and dynamic change during treatment on outcomes of patients with extremity osteosarcoma. METHODS: We retrospectively analysed 133 patients with Enneking stage IIB extremity osteosarcoma who underwent standard treatments, including univariate and multivariate analyses of patient charateritics, Hb concentrations and changes during pretreatment, neoadjuvant, adjuvant chemotherapy, and decreased Hb levels (ΔHb) to assess their prognostic value in 5-year overall survival (OS) and lung metastasis-free survival (LMFS). RESULTS: Five-year OS or LMFS were similar between patients who were anaemic and non-anaemic during pretreatment, neoadjuvant or adjuvant chemotherapy. Patients with continuously decreasing Hb had lower 5-year OS (52.3%) than those without continuous Hb decrease (68.5%, P = 0.04). Patients with ΔHb > 7.6 g/L had lower 5-year OS (57.5%) than those with ΔHb ≤7.6 g/L (75.8%, P = 0.04). However, continuous Hb decrease had no prognostic effect on 5-year LMFS. Subgroup analyses showed that patients who were anaemic during pretreatment, neoadjuvant, or adjuvant chemotherapy with ΔHb ≤7.6 g/L had better outcomes than those with ΔHb > 7.6 g/L (P < 0.05, for both). CONCLUSION: Dynamic Hb decrease and ΔHb > 7.6 predicted poor5-year OS in patients with Enneking stage IIB extremity osteosarcoma. Attempts to correct anaemia and their effects on outcomes for osteosarcoma patients should be investigated in future trials.
Subject(s)
Extremities/pathology , Hemoglobins/metabolism , Osteosarcoma/drug therapy , Adolescent , Child , Child, Preschool , Disease-Free Survival , Female , Femur/physiopathology , Humans , Male , Neoplasm Metastasis , Neoplasm Staging , Osteosarcoma/blood , Osteosarcoma/pathology , Radius/physiopathology , Tibia/physiopathology , Treatment OutcomeABSTRACT
The Retinoblastoma 1 (RB1) tumor suppressor, a member of the Retinoblastoma gene family, functions as a pocket protein for the functional binding of E2F transcription factors. About 1/3 of retinoblastoma patients harbor a germline RB1 mutation or deletion, leading to the development of retinoblastoma. Here, we demonstrate generation of a heterozygous deletion of the RB1 gene in the H1 human embryonic stem cell line using CRISPR/Cas9 nickase genome editing. The RB1 heterozygous knockout H1 cell line shows a normal karyotype, maintains a pluripotent state, and is capable of differentiation to the three germline layers.